Cyclic nucleotides have been measured in guinea pig bronchial and tracheal smooth muscles. Basal levels of the nucleotides are 0.3 plus or minus 0.5 pM/mg protein of cyclic AMP. Isoproterenol elevates cyclic AMP in both tissues within 3 minutes at concentrations of 10 to the minus 6th power thru 10 the minus 4th power M. The maximal effect is a 3.5 fold increase in the nucleotide. The beta 2 stimulant erythroprocaterol has a similar effect but neither salbutamol nor terbutaline (up to 10 to the minus 3rd power M.) alter cyclic nucleotide concentration. Isobutyl methylxanthine (IBMX) increases cyclic AMP concentrations 10-20 fold and interacts at least additively with isoproterenol or erythroprocaterol. Even in the presence of IBMX, there were no apparent increases in cyclic AMP levels exposed to terbutaline or salbutamol. In tissues tachyphylactic to 1-isoproterenol the agonist still elevated cyclic AMP levels 5 fold. Antiinflammatory drugs did not affect cyclic nucleotide levels in tracheal or bronchial tissues. Since concentration of Beta 2 agonists as low as 10 to the minus 9 power M are effective on relaxing these tissues, we do not believe that cyclic AMP is the second messenger involved in airway smooth muscle relaxation.